Ultrastructural 3D investigations of cells and cell organelles
Guenther Zellnig, Institute of Plant Sciences, University of Graz, Austria.
Ultrastructural investigations by transmission electron microscopy (TEM) are commonly performed using a limited number of ultrathin sections. In many cases the obtained results are sufficient and accurate in order to achieve a detailed characterization of cell structures on a high level of resolution.
Due to the thickness of the sections (50-80nm) only very small parts of cells and organelles can be investigated, leading to more or less two dimensional information of the investigated structures. In addition calculations of areas or volumes are neither possible nor accurate. Therefore three dimensional (3D) investigations are essential to get detailed information about the arrangement, association and the amount of subcellular structures inside a cell or organelle.
The method presented here is based on TEM serial sections and computer assisted reconstructions, which allows to obtain both morphometric data (areas and volumes) and 3D images on a high level of resolution thus enabling the detection of organelle characteristics or alterations inside cells and organelles. This 3D reconstruction method allows the visualization of a cell volume of about 3000µm3 enabling investigations of the 3D integrity of several organelles, including for instance chloroplasts with volumes of 30-40µm3, or even complete small cells like yeast cells.
Therefore this method is advantageous when compared to electron tomography which usually enables 3D visualizations of cell volumes up to 25µm3. With the 3D reconstruction and quantification technique applied here both organelle organization and quantification of their associations, being highly relevant for the intracellular translocation and communication, were evaluated in whole yeast cells.
In another study covering total plant cell organelles (chloroplasts, mitochondria) significant quantitative differences were demonstrated when compared with organelles from drought stressed plants.
